Native Receptor Research Articles

Super Resolution Dual-Energy Cone-Beam CT Imaging With Dual-Layer Flat-Panel Detector.

In flat-panel detector (FPD) based cone-beam computed tomography (CBCT) imaging, the native receptor array is usually binned into a smaller matrix size. By doing so, the signal readout speed could be increased by 4-9 times at the expense of a spatial resolution loss of 50%-67%. Clearly, such manipulation poses a key bottleneck in generating high spatial and high temporal resolution CBCT images at the same time. In addition, the conventional FPD is also difficult in generating dual-energy CBCT images. In this paper, we propose an innovative super resolution dual-energy CBCT imaging method, named as suRi, based on dual-layer FPD (DL-FPD) to overcome these aforementioned difficulties at once. With suRi, specifically, a 1D or 2D sub-pixel (half pixel in this study) shifted binning is applied instead of the conventionally aligned binning to double the spatial sampling rate during the dual-energy data acquisition. As a result, the suRi approach provides a new strategy to enable high spatial resolution CBCT imaging while at high readout speed. Moreover, a penalized likelihood material decomposition algorithm is developed to directly reconstruct the high resolution bases from these dual-energy CBCT projections containing sub-pixel shifts. Numerical and physical experiments are performed to validate this newly developed suRi method with phantoms and biological specimen. Results demonstrate that suRi can significantly improve the spatial resolution of the CBCT image. We believe this developed suRi method would greatly enhance the imaging performance of the DL-FPD based dual-energy CBCT systems in future.

Chimeric antigens displaying GPR65 extracellular loops on a soluble scaffold enabled the discovery of antibodies, which recognized native receptor

ABSTRACT GPR65 is a proton-sensing G-protein coupled receptor associated with multiple immune-mediated inflammatory diseases, whose function is relatively poorly understood. With few reagents commercially available to probe the biology of receptor, generation of an anti-GPR65 monoclonal antibody was desired. Using soluble chimeric scaffolds, such as ApoE3, displaying the extracellular loops of GPR65, together with established phage display technology, native GPR65 loop-specific antibodies were identified. Phage-derived loop-binding antibodies recognized the wild-type native receptor to which they had not previously been exposed, generating confidence in the use of chimeric soluble proteins to act as efficient surrogates for membrane protein extracellular loop antigens. This technique provides promise for the rational design of chimeric antigens in facilitating the discovery of specific antibodies to GPCRs.

Vitamin D3 administration alleviates native immune receptor, T-cell receptor beta subunit expression in a mouse model of Huntington’s disease

Vitamin D3 administration alleviates native immune receptor, T-cell receptor beta subunit expression in a mouse model of Huntington’s disease

Shedding Light on the D1 -Like Receptors: A Fluorescence-Based Toolbox for Visualization of the D1 and D5 Receptors.

Dopamine D1 -like receptors are the most abundant type of dopamine receptors in the central nervous system and, even after decades of discovery, still highly interesting for the study of neurological diseases. We herein describe the synthesis of a new set of fluorescent ligands, structurally derived from D1 R antagonist SCH-23390 and labeled with two different fluorescent dyes, as tool compounds for the visualization of D1 -like receptors. Pharmacological characterization in radioligand binding studies identified UR-NR435 (25) as a high-affinity ligand for D1 -like receptors (pKi (D1 R)=8.34, pKi (D5 R)=7.62) with excellent selectivity towards D2 -like receptors. Compound 25 proved to be a neutral antagonist at the D1 R and D5 R in a Gs heterotrimer dissociation assay, an important feature to avoid receptor internalization and degradation when working with whole cells. The neutral antagonist 25 displayed rapid association and complete dissociation to the D1 R in kinetic binding studies using confocal microscopy verifying its applicability for fluorescence microscopy. Moreover, molecular brightness studies determined a single-digit nanomolar binding affinity of the ligand, which was in good agreement with radioligand binding data. For this reason, this fluorescent ligand is a useful tool for a sophisticated characterization of native D1 receptors in a variety of experimental setups.

Constitutive Interleukin-7 Cytokine Signaling Enhances the Persistence of Epstein-Barr Virus-Specific T-Cells.

The efficacy of therapeutic T-cells is limited by a lack of positive signals and excess inhibitory signaling in tumor microenvironments. We previously showed that a constitutively active IL7 receptor (C7R) enhanced the persistence, expansion, and anti-tumor activity of T-cells expressing chimeric antigen receptors (CARs), and C7R-modified GD2.CAR T-cells are currently undergoing clinical trials. To determine if the C7R could also enhance the activity of T-cells recognizing tumors via their native T-cell receptors (TCRs), we evaluated its effects in Epstein-Barr virus (EBV)-specific T-cells (EBVSTs) that have produced clinical benefits in patients with EBV-associated malignancies. EBVSTs were generated by stimulation of peripheral blood T-cells with overlapping peptide libraries spanning the EBV lymphoma antigens, LMP1, LMP2, and EBNA 1, followed by retroviral vector transduction to express the C7R. The C7R increased STAT5 signaling in EBVSTs and enhanced their expansion over 30 days of culture in the presence or absence of exogenous cytokines. C7R-EBVSTs maintained EBV antigen specificity but were dependent on TCR stimulation for continued expansion. C7R-EBVSTs produced more rapid lymphoma control in a murine xenograft model than unmodified EBVSTs and persisted for longer. The findings have led to a clinical trial, evaluating C7R-EBVSTs for the treatment of refractory or relapsed EBV-positive lymphoma (NCT04664179).

Pseudocannabinoid H4CBD improves glucose response during advanced metabolic syndrome in OLETF rats independent of increase in insulin signaling proteins.

Cannabidiol (CBD) use has grown exponentially more popular in the last two decades, particularly among older adults (>55 yr), though very little is known about the effects of CBD use during age-associated metabolic dysfunction. In addition, synthetic analogues of CBD have generated great interest because they can offer a chemically pure product, which is free of plant-associated contaminants. To assess the effects of a synthetic analogue of CBD (H4CBD) on advanced metabolic dysfunction, a cohort of 41-wk-old Otsuka Long-Evans Tokushima Fatty (OLETF) rats were administered 200 mg H4CBD/kg by oral gavage for 4 wk. Animals were fed ad libitum and monitored alongside vehicle-treated OLETF and Long-Evans Tokushima Otsuka (LETO) rats, the lean-strain controls. An oral glucose-tolerance test (oGTT) was performed after 4 wk of treatment. When compared with vehicle-treated, OLETF rats, H4CBD decreased body mass (BM) by 15%, which was attributed to a significant loss in abdominal fat. H4CBD reduced glucose response (AUCglucose) by 29% (P < 0.001) and insulin resistance index (IRI) by 25% (P < 0.05) compared with OLETF rats. However, H4CBD did not statically reduce fasting blood glucose or plasma insulin, despite compensatory increases in skeletal muscle native insulin receptor (IR) protein expression (54%; P < 0.05). H4CBD reduced circulating adiponectin (40%; P < 0.05) and leptin (47%; P < 0.05) and increased ghrelin (75%; P < 0.01) compared with OLETF. Taken together, a chronic, high dose of H4CBD may improve glucose response, independent of static changes in insulin signaling, and these effects are likely a benefit of the profound loss of visceral adiposity.NEW & NOTEWORTHY Cannabis product use has grown in the last two decades despite the lack of research on Cannabidiol (CBD)-mediated effects on metabolism. Here, we provide seminal data on CBD effects during age-associated metabolic dysfunction. We gave 41-wk-old OLETF rats 200 mg H4CBD/kg by mouth for 4 wk and noted a high dose of H4CBD may improve glucose response, independent of static changes in insulin signaling, and these effects are likely a benefit of loss of visceral adiposity.

Mechanisms of NMDA Receptor Inhibition by Sepimostat-Comparison with Nafamostat and Diarylamidine Compounds.

N-methyl-D-aspartate (NMDA) receptors are inhibited by many amidine and guanidine compounds. In this work, we studied the mechanisms of their inhibition by sepimostat-an amidine-containing serine protease inhibitor with neuroprotective properties. Sepimostat inhibited native NMDA receptors in rat hippocampal CA1 pyramidal neurons with IC50 of 3.5 ± 0.3 µM at -80 mV holding voltage. It demonstrated complex voltage dependence with voltage-independent and voltage-dependent components, suggesting the presence of shallow and deep binding sites. At -80 mV holding voltage, the voltage-dependent component dominates, and we observed pronounced tail currents and overshoots evidencing a "foot-in-the-door" open channel block. At depolarized voltages, the voltage-independent inhibition by sepimostat was significantly attenuated by the increase of agonist concentration. However, the voltage-independent inhibition was non-competitive. We further compared the mechanisms of the action of sepimostat with those of structurally-related amidine and guanidine compounds-nafamostat, gabexate, furamidine, pentamidine, diminazene, and DAPI-investigated previously. The action of all these compounds can be described by the two-component mechanism. All compounds demonstrated similar affinity to the shallow site, which is responsible for the voltage-independent inhibition, with binding constants in the range of 3-30 µM. In contrast, affinities to the deep site differed dramatically, with nafamostat, furamidine, and pentamidine being much more active.

Cryo-EM structures reveal native GABAA receptor assemblies and pharmacology.

Type A γ-aminobutyric acid receptors (GABAARs) are the principal inhibitory receptors in the brain and the target of a wide range of clinical agents, including anaesthetics, sedatives, hypnotics and antidepressants1-3. However, our understanding of GABAAR pharmacology has been hindered by the vast number of pentameric assemblies that can be derived from 19 different subunits4 and the lack of structural knowledge of clinically relevant receptors. Here, we isolate native murine GABAAR assemblies containing the widely expressed α1 subunit and elucidate their structures in complex with drugs used to treat insomnia (zolpidem(ZOL) and flurazepam) and postpartum depression (the neurosteroid allopregnanolone(APG)). Using cryo-electron microscopy (cryo-EM)analysis and single-molecule photobleaching experiments, we uncover three major structural populations in the brain: the canonical α1β2γ2 receptor containing two α1 subunits, and two assemblies containing one α1 and either an α2 or α3 subunit, in which the single α1-containing receptors feature a more compact arrangement between the transmembrane and extracellular domains. Interestingly, APG is bound at the transmembrane α/β subunit interface, even when not added to the sample, revealing an important role for endogenous neurosteroids in modulating native GABAARs. Together with structurally engaged lipids, neurosteroids produce global conformational changes throughout the receptor that modify the ion channel pore and the binding sites for GABA and insomnia medications. Our data reveal the major α1-containing GABAAR assemblies, bound with endogenous neurosteroid, thus defining a structural landscape from which subtype-specific drugs can be developed.

Phenotypic screening of signaling motifs that efficiently induce cell proliferation

Since cell proliferation is one of the fundamental cell fates, artificial control of cell proliferation based on a receptor-engineering approach is increasingly important in therapeutic and industrial applications. Since the signal transduction properties of cytokine receptors are greatly influenced by the amino acid sequence of tyrosine motifs, here we develop a phenotypic screening approach that can directly select cell proliferation-inducing tyrosine motifs from a synthetic library. In the tyrosine motif library, amino acid sequences around the tyrosine are randomized to attain diverse binding patterns of signaling molecules. Theoretically, engineered receptors with distinct tyrosine motifs would activate signaling molecules in diverse patterns. Thus, we investigated whether tyrosine motif sequences capable of inducing cell proliferation could be selected from the cellular library expressing the motif-engineered receptors. Consequently, the selected motifs induced similar levels of cell proliferation compared to the cytoplasmic signaling domain of a native receptor. The motif-screening system was applicable to cells that may differentiate or proliferate depending on cytokine signals. To our best knowledge, this is the first report demonstrating phenotypic screening of tyrosine motifs in living cells. Our approach would open up new possibilities in the field of artificial control of cell fate based on signal transduction engineering.

Pharmacological Differences between Native Homomeric Transient Receptor Potential Canonical Type 4 Channels and Heteromeric Transient Receptor Potential Canonical Type 1/4 Channels in Lateral Septal Neurons.

Given the unique expression patterns and revelations of its critical involvement in a host of neurological disorders, the TRPC1/4/5 subgroup has become an intense target of drug development, and some compounds are now in clinical trials. However, little is known about the exact subunit composition of this subfamily of TRPC channels in various native tissues, and whether it has functional and pharmacological implications. In this study, we investigated the effects of two TRPC4 modulators located in the lateral septum, in which a metabotropic glutamate receptor (mGluR) agonist-induced plateau potential is mediated by TRPC channels composed of TRPC1 and TRPC4. Lateral septal neurons were recorded intracellularly in brain slices using sharp electrodes. Drugs were applied via bath superfusion. We showed that the plateau potential in mice lacking TRPC1 is modulated by ML204 and La3+ in a manner that is like homomeric TRPC4 channels in artificial expression systems. However, the plateau potential that is primarily mediated by heteromeric TRPC1/4 channels in lateral septal neurons in wildtype mice was modulated differently by ML204 and La3+. Our data suggest that native homomeric TRPC4 channels and heteromeric TRPC1/4 channels are pharmacologically distinct, and the current drug development strategy regarding TRPC1/4/5 may need to be reevaluated.

Affinity Maturated Transferrin Receptor Apical Domain Blocks Machupo Virus Glycoprotein Binding

Transferrin receptor 1 (TfR) delivers iron across cellular membranes by shuttling the ion carrier protein transferrin. This ability to deliver large protein ligands inside cells is taken advantage of by pathogens to infiltrate human cells. Notably, the receptor’s outermost ectodomain, the apical domain, is used as a point of attachment for several viruses including hemorrhagic arenaviruses. To better understand interactions with the receptor it would be advantageous to probe sequence determinants in the apical domain with viral spike proteins. Here, we carried out affinity maturation of our computationally designed apical domain from human TfR to identify underlying driving forces that lead to better binding. The improved variants were confirmed by in vitro surface plasmon resonance measurements with dissociation constants obtained in the lower nanomolar range. It was found that the strong binding affinities for the optimized variants matched the strength of interactions with the native receptor. The structure of the best variant was determined experimentally indicating that the conformational change in the hairpin binding motif at the protein–protein interface plays a crucial role. The experimental methodology can be straightforwardly applied to other arenavirus or pathogens that use the apical domain. It can further be useful to probe host–virus compatibility or therapeutic strategies based on the transferrin receptor decoys.

The atypical 'hippocampal' glutamate receptor coupled to phospholipase D that controls stretch-sensitivity in primary mechanosensory nerve endings is homomeric purely metabotropic GluK2.

A metabotropic glutamate receptor coupled to phospholipase D (PLD-mGluR) was discovered in the hippocampus over three decades ago. Its pharmacology and direct linkage to PLD activation are well established and indicate it is a highly atypical glutamate receptor. A receptor with the same pharmacology is present in spindle primary sensory terminals where its blockade can totally abolish, and its activation can double, the normal stretch-evoked firing. We report here the first identification of this PLD-mGluR protein, by capitalizing on its expression in primary mechanosensory terminals, developing an enriched source, pharmacological profiling to identify an optimal ligand, and then functionalizing it as a molecular tool. Evidence from immunofluorescence, western and far-western blotting indicates PLD-mGluR is homomeric GluK2, since GluK2 is the only glutamate receptor protein/receptor subunit present in spindle mechanosensory terminals. Its expression was also found in the lanceolate palisade ending of hair follicle, also known to contain the PLD-mGluR. Finally, in a mouse model with ionotropic function ablated in the GluK2 subunit, spindle glutamatergic responses were still present, confirming it acts purely metabotropically. We conclude the PLD-mGluR is a homomeric GluK2 kainate receptor signalling purely metabotropically and it is common to other, perhaps all, primary mechanosensory endings.

Ligand-Dependent Intramolecular Motion of Native Nicotinic Acetylcholine Receptors Determined in Living Myotube Cells via Diffracted X-ray Tracking.

Nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels that play an important role in signal transduction at the neuromuscular junction (NMJ). Movement of the nAChR extracellular domain following agonist binding induces conformational changes in the extracellular domain, which in turn affects the transmembrane domain and opens the ion channel. It is known that the surrounding environment, such as the presence of specific lipids and proteins, affects nAChR function. Diffracted X-ray tracking (DXT) facilitates measurement of the intermolecular motions of receptors on the cell membranes of living cells, including all the components involved in receptor function. In this study, the intramolecular motion of the extracellular domain of native nAChR proteins in living myotube cells was analyzed using DXT for the first time. We revealed that the motion of the extracellular domain in the presence of an agonist (e.g., carbamylcholine, CCh) was restricted by an antagonist (i.e., alpha-bungarotoxin, BGT).

Modeling of ACE2 and antibodies bound to SARS-CoV-2 provides insights into infectivity and immune evasion.

Given the COVID-19 pandemic, there is interest in understanding ligand-receptor features and targeted antibody-binding attributes against emerging SARS-CoV-2 variants. Here, we developed a large-scale structure-based pipeline for analysis of protein-protein interactions regulating SARS-CoV-2 immune evasion. First, we generated computed structural models of the Spike protein of 3 SARS-CoV-2 variants (B.1.1.529, BA.2.12.1, and BA.5) bound either to a native receptor (ACE2) or to a large panel of targeted ligands (n = 282), which included neutralizing or therapeutic monoclonal antibodies. Moreover, by using the Barnes classification, we noted an overall loss of interfacial interactions (with gain of new interactions in certain cases) at the receptor-binding domain (RBD) mediated by substituted residues for neutralizing complexes in classes 1 and 2, whereas less destabilization was observed for classes 3 and 4. Finally, an experimental validation of predicted weakened therapeutic antibody binding was performed in a cell-based assay. Compared with the original Omicron variant (B.1.1.529), derivative variants featured progressive destabilization of antibody-RBD interfaces mediated by a larger set of substituted residues, thereby providing a molecular basis for immune evasion. This approach and findings provide a framework for rapidly and efficiently generating structural models for SARS-CoV-2 variants bound to ligands of mechanistic and therapeutic value.

Tailoring the Inherent Properties of Biobased Nanoparticles for Nanomedicine.

Biobased nanoparticles are at the leading edge of the rapidly developing field of nanomedicine and biotherapeutics. Their unique size, shape, and biophysical properties make them attractive tools for biomedical research, including vaccination, targeted drug delivery, and immune therapy. These nanoparticles are engineered to present native cell receptors and proteins on their surfaces, providing a biomimicking camouflage for therapeutic cargo to evade rapid degradation, immune rejection, inflammation, and clearance. Despite showing promising clinical relevance, commercial implementation of these biobased nanoparticles is yet to be fully realized. In this perspective, we discuss advanced biobased nanoparticle designs used in medical applications, such as cell membrane nanoparticles, exosomes, and synthetic lipid-derived nanoparticles, and highlight their benefits and potential challenges. Moreover, we critically assess the future of preparing such particles using artificial intelligence and machine learning. These advanced computational tools will be able to predict the functional composition and behavior of the proteins and cell receptors present on the nanoparticle surfaces. With more advancement in designing new biobased nanoparticles, this field of research could play a key role in dictating the future rational design of drug transporters, thereby ultimately improving overall therapeutic outcomes.

Enhancing CAR T-cell Therapy Using Fab-Based Constitutively Heterodimeric Cytokine Receptors.

Adoptive T-cell therapy aims to achieve lasting tumor clearance, requiring enhanced engraftment and survival of the immune cells. Cytokines are paramount modulators of T-cell survival and proliferation. Cytokine receptors signal via ligand-induced dimerization, and this principle has been hijacked utilizing nonnative dimerization domains. A major limitation of current technologies resides in the absence of a module that recapitulates the natural cytokine receptor heterodimeric pairing. To circumvent this, we created a new engineered cytokine receptor able to constitutively recreate receptor-heterodimer utilizing the heterodimerization domain derived from the IgG1 antibody (dFab_CCR). We found that the signal delivered by the dFab_CCR-IL2 proficiently mimicked the cytokine receptor heterodimerization, with transcriptomic signatures like those obtained by activation of the native IL2 receptor. Moreover, we found that this dimerization structure was agnostic, efficiently activating signaling through four cytokine receptor families. Using a combination of in vivo and in vitro screening approaches, we characterized a library of 18 dFab_CCRs coexpressed with a clinically relevant solid tumor-specific GD2-specific chimeric antigen receptor (CAR). Based on this characterization, we suggest that the coexpression of either the common β-chain GMCSF or the IL18 dFab_CCRs is optimal to improve CAR T-cell expansion, engraftment, and efficacy. Our results demonstrate how Fab dimerization is efficient and versatile in recapitulating a cytokine receptor heterodimerization signal. This module could be applied for the enhancement of adoptive T-cell therapies, as well as therapies based on other immune cell types. Furthermore, these results provide a choice of cytokine signal to incorporate with adoptive T-cell therapies.

A method for polyclonal antigen-specific T cell-targeted genome editing (TarGET) for adoptive cell transfer applications

Adoptive cell therapy of donor-derived, antigen-specific Tcells expressing native Tcell receptors (TCRs) is a powerful strategy to fight viral infections in immunocompromised patients. Determining the fate of Tcells following patient infusion hinges on the ability to track them invivo. While this is possible by genetic labeling of parent cells, the applicability of this approach has been limited by the non-specificity of the edited Tcells. Here, we devised a method for CRISPR-targeted genome integration of a barcoded gene into Epstein-Barr virus-antigen-stimulated Tcells and demonstrated its use for exclusively identifying expanded virus-specific cell lineages. Our method facilitated the enrichment of antigen-specific Tcells, which then mediated improved cytotoxicity against Epstein-Barr virus-transformed target cells. Single-cell and deep sequencing for lineage tracing revealed the expansion profile of specific Tcell clones and their corresponding gene expression signature. This approach has the potential to enhance the traceability and the monitoring capabilities during immunotherapeutic Tcell regimens.

Optical tools for visualizing and controlling human GLP-1 receptor activation with high spatiotemporal resolution

The glucagon-like peptide-1 receptor (GLP1R) is a broadly expressed target of peptide hormones with essential roles in energy and glucose homeostasis, as well as of the blockbuster weight-loss drugs semaglutide and liraglutide. Despite its large clinical relevance, tools to investigate the precise activation dynamics of this receptor with high spatiotemporal resolution are limited. Here, we introduce a novel genetically encoded sensor based on the engineering of a circularly permuted green fluorescent protein into the human GLP1R, named GLPLight1. We demonstrate that fluorescence signal from GLPLight1 accurately reports the expected receptor conformational activation in response to pharmacological ligands with high sensitivity (max ΔF/F0=528%) and temporal resolution (τON = 4.7 s). We further demonstrated that GLPLight1 shows comparable responses to glucagon-like peptide-1 (GLP-1) derivatives as observed for the native receptor. Using GLPLight1, we established an all-optical assay to characterize a novel photocaged GLP-1 derivative (photo-GLP1) and to demonstrate optical control of GLP1R activation. Thus, the new all-optical toolkit introduced here enhances our ability to study GLP1R activation with high spatiotemporal resolution.

Optical tools for visualizing and controlling human GLP-1 receptor activation with high spatiotemporal resolution.

The glucagon-like peptide-1 receptor (GLP1R) is a broadly expressed target of peptide hormones with essential roles in energy and glucose homeostasis, as well as of the blockbuster weight-loss drugs semaglutide and liraglutide. Despite its large clinical relevance, tools to investigate the precise activation dynamics of this receptor with high spatiotemporal resolution are limited. Here, we introduce a novel genetically encoded sensor based on the engineering of a circularly permuted green fluorescent protein into the human GLP1R, named GLPLight1. We demonstrate that fluorescence signal from GLPLight1 accurately reports the expected receptor conformational activation in response to pharmacological ligands with high sensitivity (max ΔF/F0=528%) and temporal resolution (τON = 4.7 s). We further demonstrated that GLPLight1 shows comparable responses to glucagon-like peptide-1 (GLP-1) derivatives as observed for the native receptor. Using GLPLight1, we established an all-optical assay to characterize a novel photocaged GLP-1 derivative (photo-GLP1) and to demonstrate optical control of GLP1R activation. Thus, the new all-optical toolkit introduced here enhances our ability to study GLP1R activation with high spatiotemporal resolution.

Computational study unravels inhibitory potential of epicatechin gallate against inflammatory and pyroptosis‐associated mediators in COVID‐19

AbstractCoronavirus disease‐19 (COVID‐19) is the global health emergency caused by SARS‐CoV‐2. Upon infection, antigenic determinants of the virus trigger massive production of proinflammatory/pyroptosis‐associated proteins, resulting in cytokine storm, tissue damage, and multiorgan failure. Therefore, these proinflammatory/pyroptosis‐associated mediators are promising therapeutic targets to combat COVID‐19. Epicatechin gallate (ECG) is a polyphenol found in green tea. It has antioxidative and anti‐inflammatory properties. Hence, in the present study, ECG was selected to explore its binding potential for inflammatory mediators such as interleukins, interferon‐γ (IFNγ), and tumor necrosis factor‐α (TNF‐α), along with their native receptors. In addition, the interacting potential of ECG with pyroptosis‐associated proteins, viz. caspases and BAX has also been investigated. Molecular docking analysis has revealed that ECG interacts with interleukins, IFNγ, TNF‐α, cytokine receptors, caspase‐1/4/11, and BAX with significant binding affinity. Several amino acid residues of these mediators were blocked by ECG through stable hydrogen bonds and hydrophobic contacts. ECG interacted with caspase‐11, BAX, and TNF‐R1 with better binding affinities. Therefore, the present in silico study indicates that ECG could be a potential drug to subvert cytokine storm and pyroptosis during COVID‐19.